Phosphorus is one of the most important nutrients for plants, being essential for their growth and a structural component of nucleic acids, phospholipids, intermediary metabolites and numerous other biological molecules. In plants, the only readily absorbed form of exogenous phosphorus is inorganic phosphate (P.sub.i) (Bieleski, 1973). When the amount of available phosphate is low, plants are unable to grow vigorously and productively. When phosphate is absent, growth is halted and the plant dies.
The phosphate to nitrogen ratio in plants affects both the temporal and quantitative characteristics of flowering (Salisbury and Ross, 1985). Relatively high phosphate advances maturity in plants, whereas relatively low phosphate results in little or no flowering taking place. Phosphate levels are also known to affect the biomass ratio between root and shoot. Specifically, phosphate deprivation causes preferential growth of roots (Lefebvre et al., 1982). Thus, in many environments, the availability of phosphorus becomes a major factor limiting the growth and reproduction of photosynthetic organisms.
Plants and other photosynthetic organisms are either sessile or restricted in movement and therefore limited in nutrient availability to their immediate environment. As a result, photosynthetic organisms require signal transduction pathways in order to trigger cellular responses to adverse environmental stimuli. As part of the adenosine nucleotides, ADP and ATP, which are the currency of cellular energy, phosphorus is critical to bioenergetics. The covalent addition or removal of a phosphate group to or from a biological substrate (phosphorylation and dephosphorylation, respectively) often functions as a kind of regulatory "on/off switch" in cellular metabolism and signal transduction. For example, the phosphorylation and dephosphorylation of certain membrane-bound receptor protein kinases and their substrates are key to various signal transduction pathways, including pathways of plant hormones such as ethylene (Kieber et al., 1993) and abscisic acid (Anderberg and Walker-Simmons, 1992). Self-incompatibility with respect to pollination and fertilization also involves the activity of protein kinases encoded by S-locus genes (Tantikanjana et al., 1993; Zhang and Walker, 1993). Regulatory signals can also trigger mechanisms for disease and pest control.
Although the nature of the phosphate-starvation response has been investigated in plants, little is known of the molecular mechanisms that regulate phosphorus uptake and metabolism. Few genes associated with phosphate-starvation expression in plants have been identified and isolated. To date, no reports of promoters which specifically respond to phosphate-starvation conditions are known.
Among the promoter sequences available for the genetic engineering of plants, depending upon the transcription initiation characteristics desired, (strength, tissue specificity, developmental specificity, etc.), different promoters can be employed to initiate transcription of a DNA sequence of interest joined at the 3' end of the promoter region. For example, promoters such as the 35S Cauliflower Mosaic Virus (CaMV 35S), mannopine synthase (mas) and octopine synthase (ocs) have been used successfully to direct the expression of desired nucleic acid sequences in transformed plant tissue. When expressed in a transgenic plant, DNA sequences under the control of these promoters are found at relatively low or moderate levels and are expressed fairly evenly (i.e. constitutively) throughout the plant. See, for example, van der Zaal, et al. (1991) Plant Mol. Biol. 16:983; Ohl, et al. (1990) Cell 2:837.
Of particular interest, however, are promoters which demonstrate enhanced transcription initiation characteristics in rapidly dividing cells or rapidly growing tissue, against stress or other detrimental factors. Likewise, there are advantages with the increased expression of a DNA sequence of interest under the regulation of an inducible promoter. Such promoters regulate the expression of genes in response to environmental factors, such as light, wounding, exposure to heavy metals, low nutrient status, and/or temperature. However, isolated DNA elements responsive to environmental nutrients are rare.
The discovery of new promoters with useful transcript initiation patterns, especially ones having very strong promoter activity, are essential for the controlled expression of desirable nucleic acid sequences. Promoters which show enhanced activity induced by environmental phenomena are also of special interest for many genetic engineering tools to enhance plant characteristics generally, as these critical gene sequences are abundant in young or stressed plant tissue. Thus, a need exists for promoter sequences which can be used in recombinant DNA constructions to enable the external control of the expression of genes which can confer agronomic advantages when expressed at the proper time.